Synthesis and stereoselective κ-receptor binding of methylated analogues of GR-89.696

Article abstract of DOI:10.1016/S0223-5234(00)01208-3

Stereoselective synthesis of all four stereoisomers of methylated analogues 8 of the κ-receptor agonist GR-89.696 is presented. Starting with orthogonally protected piperazine derivatives (R,R)-4 and (S,S)-4, the reaction sequence involves oxidation, redu

Full text of DOI:10.1016/S0223-5234(00)01208-3

Eur. J. Med. Chem. 36 (2001) 211–214
´
© 2001 Editions scientifiques et me´dicales Elsevier SAS. All rights reserved
PII: S0223-5234(00)01208-3/SCO
Short communication
Synthesis and stereoselective k-receptor binding of methylated analogues of
GR-89.696
Caroline Ro¨hra, Stella Soukarab, Bernhard Wu¨nschb*
aInstitut fu¨r Anorganische und Analytische Chemie der Universita¨t Freiburg, Albertstraße 21, 79104 Freiburg, Germany
bPharmazeutisches Institut der Universita¨t Freiburg, Hermann-Herder-Straße 9, 79104 Freiburg i. Br., Germany
Received 28 August 2000; revised 30 November 2000; accepted 1 December 2000
Dedicated to Prof. Dr. August W. Frahm on the occasion of his 65th birthday
Abstract – Stereoselective synthesis of all four stereoisomers of methylated analogues 8 of the k-receptor agonist GR-89.696 is
presented. Starting with orthogonally protected piperazine derivatives (R,R)-4 and (S,S)-4, the reaction sequence involves oxidation,
reductive amination and modification of the piperazine nitrogen protective groups. The configuration of the stereocentre in
a-position to the pyrrolidine moiety is determined by X-ray structure analysis of (R,S)-8. In receptor-binding studies with the
radioligand U-69.593, the stereoisomer with (S)-configuration at both stereogenic centres (S,S)-8 displayed the highest k-receptor
´
affinity with a Ki-value of 0.67 nM. © 2001 Editions scientifiques et me´dicales Elsevier SAS
k-receptor agonists / 2-[1-(pyrrolidin-1-yl)ethyl]-piperazines / diastereoselective synthesis / X-ray crystal structure / stereoselective
k-receptor binding
1. Introduction
tive k-receptor agonists. In the diamide series, the
N1-propionyl-substituted piperazine 1a revealed the
In the brain and peripheral tissues, three distinct
opioid receptor subtypes, m, k and d, have been iden-
tified. It is well established that activation of each of
these receptor subtypes can produce analgesic effects.
Among ligands for these opioid receptor subtypes,
k-agonists are of particular interest, since only mini-
mal physical dependence, respiratory depression and
obstipation are associated with a strong analgesic
profile. Apart from their analgesic effects, k-agonists
may play a role as neuroprotectants in certain animal
models of cerebral ischaemia. However, application
of k-agonists may be limited by adverse side effects
including sedation, dysphoria and strong diuresis [1,
2].
highest k-receptor affinity in the rabbit vas deferens
functional in vitro assay (IC50 =6.9 nM). A dramatic
enhancement of the k-receptor affinity by a factor of
170 was achieved with the bioisosteric carbamate 1b
(GR-89.696; IC50 =0.041 nM), the k-receptor affinity
of which resides almost exclusively in the (R)-enan-
tiomer [(R)-1b: IC50 =0.018 nM] [3, 4] (figure 1).
The 3-(pyrrolidin-1-ylmethyl) substituted 1,4-dia-
cylpiperazines 1 belong to the most active and selec-
* Correspondence and reprints
E-mail address: wuensch@ruf.uni-freiburg.de (B. Wu¨nsch).
Figure 1.

212
C. Ro¨hr et al. / European Journal of Medicinal Chemistry 36 (2001) 211–214
The oxidation of the secondary alcohol (R,R)-4
In the 2-(aminomethyl)piperidine substance class of
succeeded with a catalytic amount of tetrapropylam-
monium perruthenate (Pr4N+ RuO4–, TPAP) and an
excess of the reoxidant N-methylmorpholine-N-oxide
(NMMO) [7, 8] to provide the 3-acetylpiperazine (R)-
5 [9] in 85% yield.
Attempts to perform the reductive amination of the
ketone (R)-5 with pyrrolidine and NaBH3CN accord-
ing to the procedure of Borch and coworkers [10]
failed. However, a variation using the Lewis acid
Ti(OiPr)4 to accelerate iminium ion formation [11] led
to the diastereomeric pyrrolidines (R,S)-6 and (R,R)-
6 in the ratio 7:3. Flash chromatographic separation
afforded the diastereomers (R,S)-6 and (R,R)-6 in 62
and 25% yield, respectively.
The last steps in the reaction sequence represent
modifications of the piperazine nitrogen protective
groups. At first, the benzyl protective group (Bn) of 6
was hydrogenolytically (H2, Pd/C) [12] removed and,
subsequently, the secondary amines were acylated
with (3,4-dichlorophenyl)acetic acid (DCPAA) in the
presence of 1,1’-carbonyldiimidazole (CDI) to yield
the diastereomeric phenylacetamides (R,S)-7 and
(R,R)-7, respectively. Finally, the tert-butoxycar-
bonyl protective group (BOC) of 7 was cleaved with
trifluoroacetic acid/CH2Cl2 [13]. The resulting crude
secondary amines were acylated with propionyl chlo-
ride and NaOH in a biphasic system to afford the
diacyl piperazines (R,S)-8 and (R,R)-8, respectively
(figure 3).
k-agonists an additional methyl group in a-position
to the dimethylamino group is, depending on the
stereochemistry, able to enhance k-receptor affinity
[5]. Accordingly, we envisioned that yet unknown
methylated (pyrrolidinylmethyl)piperidines or -piper-
azines of general structure 2 should be potent k-recep-
tor ligands. This prompted us to undertake the
synthesis and in vitro testing of those compounds.
One of the major issues of this study was to investi-
gate the influence of the stereochemistry of the second
chiral centre on k-receptor affinity.
In this communication, we report on the stereose-
lective synthesis of all four stereoisomeric methylated
(pyrrolinin-1-ylmethyl)piperazines 8, the elucidation
of their stereochemistry as well as on their k-receptor
affinities.
2. Chemistry
The synthesis of the piperazines 8 with (R)-configu-
ration in position 3 starts from the proteinogenic
amino acid (2S,3R)-threonine, which was transformed
via the intermediate bicyclic piperazinedione (S,R)-3
into the orthogonally protected (R,R)-configured 3-
(1-hydroxyethyl)piperazine (R,R)-4 according to liter-
ature [6]. During LiAlH4 reduction of the bicyclic
piperazinedione (S,R)-3 the stereodescriptor denoting
the stereocentre in position 3 is changed from (S) in 3
to (R) in the piperazines 4–8 since the priority of the
substituents according to the CIP rules is changed
(figure 2) [6].
The respective (3S)-configured enantiomers (S,R)-8
and (S,S)-8 were prepared in an analogous manner
starting with the enantiomeric amino acid (2R,3S)-
threonine.
3. X-ray analysis
The absolute configuration of the new stereocentre
generated by reductive amination of the ketone (R)-5,
could not be determined unequivocally by means of
NMR spectroscopy. However, X-ray suitable crystals
of the major diastereomer (R,S)-8 could be obtained
by means of purification by flash chromatography
and repeated recrystallisation from ethyl acetate. Ac-
cording to the X-ray structure analysis (figure 4),
unlike configurations [14] could be established un-
doubtedly for the two adjacent stereocentres. Since
the absolute configuration of the stereocentre in posi-
Figure 2.

C. Ro¨hr et al. / European Journal of Medicinal Chemistry 36 (2001) 211–214
213
tion 3 of the piperazine ring is known from the
synthetic pathway to be (R), the newly generated side
chain stereocentre has to be (S)-configured. Conclu-
sively, and since the C-3 stereocentre remains unaf-
fected in the reaction sequence from 6 to 8, the
stereochemistry of the intermediates [(R,S)-7 and
(R,S)-6)] and diastereomers [(R,R)-6, (R,R)-7, (R,R)-
8] is established beyond any doubt.
In addition to elucidation of the configuration, the
X-ray analysis indicates that the 1-(pyrrolidin-1-
yl)ethyl residue in position 3 of the piperazine moiety
adopts an axial orientation. In solution, the C-3 sub-
stituent is also most likely to be forced into the axial
position because of the allylic 1,3-strain (A1,3 strain)
of the amide moiety [16]. This result is in accordance
with the X-ray structure of piperazine and piperidine
analogues with (aminomethyl) substituents [3, 17].
Figure 3.
Figure 5.
Figure 4. ORTEP drawing of the X-ray structure of (R,S)-8 [15].

214
C. Ro¨hr et al. / European Journal of Medicinal Chemistry 36 (2001) 211–214
Table I. k-Receptoraffinities of the stereoisomeric piperazines 8.
der Chemischen Industrie. Thanks are also due to the
Degussa AG for donation of chemicals.
Compound
Ki (nM)9S.E.M.
(R,S)-8
(S,R)-8
(R,R)-8
(S,S)-8
\10 000
References
4396.1
4.290.38
0.6790.17
0.4990.16
[1] Scopes D.I.C., Drug Fut. 18 (1993) 933–947.
[2] Holzgrabe U., Nachtsheim C., Siener T., Drosihn S., Brandt
W., Pharmazie 52 (1997) 4–22.
U-50.488
[3] Naylor A., Judd D.B., Lloyd J.E., Scopes D.I.C., Hayes A.G.,
Birch P.J., J. Med. Chem. 36 (1993) 2075–2083.
[4] Birch P.J., Rogers H., Hayes A.G., Hayward N.J., Tyers M.B.,
Scopes D.I.C., Naylor A., Judd D.B., Br. J. Pharmacol. 103
(1991) 1819–1823.
4. Receptor-binding studies
[5] Vecchietti V., Giordani A., Giardina G., Colle R., Clarke G.D.,
J. Med. Chem. 34 (1991) 397–403.
In order to determine the k-receptor affinities of the
stereoisomeric piperazines 8, in vitro receptor-binding
assays were performed using guinea-pig brain mem-
brane preparations as receptor material [18]. k-Recep-
tors were labelled with the radioligand [3H]-U-69.593
and non-specific binding was defined with an excess
of U-50.488 (figure 5) [19, 20].
The Ki-values of the stereoisomeric piperazines 8,
which are summarised in table I, reveal that in this
substance class the k-receptor affinity is strongly de-
pendent on the stereochemistry.
Thus, the like-configured [14] isomers (R,R)-8 and
(S,S)-8 show much higher affinities when compared to
the unlike stereoisomers (R,S)-8 and (S,R)-8. In both
diastereomeric pairs, the enantiomers with (3S)-
configuration display higher k-receptor affinities than
their (3R)-enantiomers. This observation is in accor-
dance with the reported higher k-receptor affinity of
(R)-1b [3, 4]. [Note, that the stereochemistry in posi-
tion 3 of (S,S)-8 and (S,R)-8 is equivalent to the
stereochemistry of (R)-1b, since the priority of the
substituents according to the CIP rules is changed.]
Comparison of the k-receptor affinities of the
diastereomers (S,S)-8 (Ki =0.67 nM) and (S,R)-8
(Ki=43 nM) indicates that (S)-configuration in the
[6] S. Soukara, B. Wu¨nsch, Synthesis (1999) 1739–1746.
[7] Griffith W.P., Ley S.V., Aldrichimica Acta 23 (1990) 13–19.
[8] S.V. Ley, J. Norman, W.P. Griffith, S.P. Marsden, Synthesis
(1994) 639–666.
[9] All new compounds have been analysed by one- and two-di-
mensional 1H- and 13C-NMR spectroscopy, by IR and mass
spectroscopy, by polarimetry and elemental analyses.
[10] Borch R.F., Benstein M.D., Durst H.D., J. Am Chem. Soc. 93
(1971) 2897–2904.
[11] Mattson R.J., Pham K.M., Leuck D.J., Cowen K.A., J. Org.
Chem. 55 (1990) 2552–2554.
[12] T.W. Greene, P.G.M Wuts, in: Protective Groups in Organic
Synthesis, John Wiley & Sons: New York, Chichester, Bris-
bane, Toronto, Singapore, 2nd ed., 1991, p. 364 f.
[13] Ref. 12, pp. 327 f.
[14] E.L. Eliel, S.H. Wilen, in: Stereochemistry of Organic Com-
pounds, John Wiley & Sons: New York, Chichester, Brisbane,
Toronto, Singapore, 1994, pp. 120 f. and 1210.
[15] Crystallographic data (excluding structure factors) for the
structure in this paper has been deposited with the Cambridge
Crystallographic Data Centre as supplementary publication no.
CCDC 157285. Copies of the data can be obtained, free of
charge, on application to CCDC, 12 Union Road Cambridge,
CBZ
1 EZ, UK, (fax: +44 1223 336033 or e-mail: de-
posit@ccdc.cam.ac.uk)
[16] Seebach D., Lamatsch B., Amstutz R., Beck A.K., Dobler M.,
Egli M., Fitzi R., Gautschi M., Herradon B., Hidber P.C.,
Irwin J.J., Locher R., Maestro M., Maetzke T., Mourino A.,
Pfammatter E., Plattner D.A., Schickli C., Schweizer W.B.,
Seiler P., Stucky G., Petter W., Escalante J., Juaristi E., Quin-
tana D., Miravitlles C., Molins E., Helv. Chim. Acta 75 (1992)
913–934.
side chain increases k-receptor affinity. Thus, combi-
.
nation of (S)-configuration in the piperazine ring sys-
tem with (S)-configuration in the side chain provides
(S,S)-8 with the highest k-receptor affinity.
[17] Scopes D.I.C., Hayes N.F., Bays D.E., Belton D., Brain J.,
Brown D.S., Judd D.B., McElroy A.B., Meerholz C.A., Naylor
A., Sheehan M.J., Birch P.J., Tyers M.B., J. Med. Chem. 35
(1992) 490–501.
[18] Smith J.A.M., Hunter J.C., Hill R.G., Hughes J., J. Neu-
rochem. 53 (1989) 27–36.
Acknowledgements
[19] Wanner K.Th., Praschak I., Ho¨fner G., Beer H., Arch. Pharm.
Pharm. Med. Chem. 329 (1996) 11–22.
We gratefully acknowledge the financial support of
the Deutsche Forschungsgemeinschaft and the Fonds
[20] Archer S., Seyed-Mozaffari A., Jiang Q., Bidlack J.M., J. Med.
Chem. 37 (1994) 1578–1585.